Protein overlapping cleavage

The order in which the tryptic peptides occur must then be determined by another technique, e.g. according to a scheme of overlapping cleavages with other proteases (pepsin, ch3onotrypsin, papain, and others). The first extensive sequence of amino acids was determined on insulin (Sanger 1954), which is a polypeptide consisting of 51 amino acids. The sequential Analysis of true proteins was still beset with serious obstacles. Finally in 1959, ribonuclease vith 124 amino acid... 

MEROPS database published by Rawlings et ai., 2004). Each of these has overlapping but distinct preferences for peptide bonds and, hence, together can affect the efficient breakdown of proteins. As cysteine proteases generally operate at slightly acid pH (5.0-6.5), they are suitable for functioning within the low pH of the gut lumen (for S. mansoni) where initial cleavage steps take place (Dresden et ai., 1981 Dalton et ai., 1996 Brindley et ai., 1997 Tort et ai., 1999 Sajid and McKerrow, 2002). 

It can be concluded that the bacterial proteinases can be applied to many proplems where extensive enzymatic proteolysis is required. Because of their wide substrate specificity, they are not useful for producing one of the major sets of peptides which are required to provide overlapping sequences within large polypeptides or proteins. On the other hand, where structural studies of small peptides require extensive cleavage at a variety of bonds, the bacterial proteinases are excellent hydrolytic agents. The products of proteinase action are often quite analogous to the products... 

The peptides obtained by specific chemical or enzymatic cleavage are separated by some type of chromatography. The sequence of each purified peptide is then determined by the Edman method. At this point, the amino acid sequences of segments of the protein are known, but the order of these segments is not yet defined. How can we order the peptides to obtain the primary structure of the original protein The necessary additional information is obtained from overlap... 

The sequence of amino acids in the polypeptide chain (i.e., the primary structure of a polypeptide or protein) can be established by selective chemical and enzymatic cleavage of the protein followed by separation, amino acid analysis, and sequence determination of all peptide fragments. The entire amino acid sequence is established by overlapping identical regions of the individual fragments. The following problem illustrates the procedure. 

---